This proposal concerns the roles of cytokines in human, immune mediated (type I) diabetes (IMD). The underlying hypothesis is that intra-islet expressions of TH1 cytokines (IFN-gamma, TNF-a, IL-12), promote pancreatic cell destruction and progression to diabetes while TH2 cytokines (IL-4, IL-10, IL-13) are associated with a benign, possibly protective insulitis, and a non-progressive course. We will measure cytokines produced by: 1) peripheral blood T cells undergoing a recall response after prolonged in-vitro exposure to islet cell antigens, as restricted by HLA-DR/DQ; 2) by NK1.1+ T cells expressing Va24J Q receptors as restricted by CDId; and 3) by direct measurement in sera. The influence of cytokines on the speed of pancreatic cell destruction in the natural history of IMD as seen in autoantibody positive relatives, and their possible roles in affecting the diabetic outcomes of an antigen (insulin) based intervention trial will be studied. The patients will be those ascertained as part of an oral (insulin) tolerance trial in newly diagnosed diabetics aged between 4 and 60 years devised by the PI, and those participating in the type 1 diabetes prevention trial DPT-1. Specific Aim 1, is to examine cytokine levels in the recall responses to in-vitro immunizations of peripheral blood T cells by insulin, insulin B chain, GAD65 and IA-2 from 30+ newly diagnosed IMD patients and HLA-DR/DQ matched controls, as restricted by DR and/or DQ and CD1d antibodies. Sera from 300 newly diagnosed patients will also be studied and their cytokine profiles will be related to HLA-DR/DQ phenotypes and ages at onset of diabetes. Specific Aim 2 is to examine cytokine levels (IL-4, 13, IFN-, IL-12, TGF-) in stored sera from groups of relatives (n=100) of patients screened as part of DPT-1 who are at high risk (2 or more of ICA, GAD65A, IAA and IA-2A), low risk (one antibody +), or near no risk (no antibodies) for IMD. The results will be related to expected risk of diabetes, and to HLA-DR/DQ. Specific Aim 3 is to study the changes of cytokines in the sera, and stimulated (anti-CD3/PHA) levels of cytokine mRNAs plus tyrosine phosphorylation of the transcription factor STAT-6 induced by intrinsic IL-4/13 in sequential blood samples of newly diagnosed patients enrolled in the oral insulin tolerance trial. Time permitting, serial determinations of T cell responses to islet cell antigens should be made too. The insights gained in these studies should lead to novel means of future interventions aimed at deviating progressive (TH1 cytokines) to non-progressive (TH2 cytokines) insulitis lesions, through immunizations by islet cell antigenic peptides, long acting cytokines or both.